Mechanism of Ligand Gating in Potassium Channels

钾通道配体门控机制

• 批准号: 6930943
• 负责人: YOUXING JIANG
• 金额: $ 28.08万
• 依托单位: UT SOUTHWESTERN MEDICAL CENTER
• 依托单位国家: 美国
• 财政年份: 2004
• 资助国家: 美国
• 起止时间: 2004-09-01 至 2009-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6930943
• 关键词: Archaea; X ray crystallography; binding sites; bioenergetics; calcium ion; chemical binding; conformation; electrophysiology; genetic manipulation; ligands; potassium channel; protein protein interaction; protein structure; protein structure function; structural biology

DESCRIPTION (provided by applicant): Potassium channels control the flow of K+ into and out of the cell, and are ubiquitously expressed in nearly all organisms ranging from the simplest bacterium to humans. One of the most important properties of K+ channels is gating, that is, the opening and closing of the channel in response to external stimuli. K+ channel gating plays a vital role in many important biological processes such as the excitation of nerve and muscle cells. Understanding K+ channel gating will provide basic, fundamental knowledge about K+ channel-related biological activities and diseases. Currently, there is a large body of functional data on K+ channel gating activity, but little is known about the structure that underlies the gating process. The broad goal of my research is to understand the structure and mechanics of the K+ channel. More specifically, our lab will focus on studying the gating mechanism in what has already proven to be an excellent model system, a Ca2+-activated K+ channel called MthK, from the archaebacterium Methanobacterium thermoautotrophicum. Our approach will be multi-disciplinary, utilizing both X-ray crystallography and electrophysiology. The proposed research has three specific aims. The first specific aim is to determine the X-ray structure of the MthK channel in a closed conformation. This combined with the known structure of MthK in the open form will provide the first example of detailed, atomic resolution pictures of an ion channel in both the opened and closed conformations. The second specific aim is to obtain high resolution X-ray structures of MthK with the help of monoclonal antibodies. The high-resolution structures will elucidate atomic details of the specific interactions at the ligand binding site and the protein-protein contacts that underlie conformational changes. The third specific aim is to study the functional mechanics of the coupling between Ca2+ binding and channel gating. We will use structure-based mutagenesis combined with single channel electrophysiological recordings to analyze the energetic process of ligand gating.

描述（由申请人提供）：钾通道控制K+进入细胞的流动，并且在几乎所有的生物中都无处不在，从最简单的细菌到人类。 K+通道最重要的特性之一是门控，即响应外部刺激的通道的开口和关闭。 K+通道门控在许多重要的生物学过程中起着至关重要的作用，例如神经和肌肉细胞的激发。了解K+通道门控将提供有关K+通道相关的生物学活动和疾病的基本基础知识。当前，关于K+通道门控活动的功能数据大量，但对基础门控过程的结构知之甚少。我的研究的广泛目标是了解K+通道的结构和机制。更具体地说，我们的实验室将专注于研究已经证明是出色的模型系统的门控机制，即来自甲虫菌群嗜热菌的Ca2+激活的K+通道，称为MTHK，称为MTHK。我们的方法将是多学科的，利用X射线晶体学和电生理学。拟议的研究具有三个具体目标。第一个具体目的是在封闭构象中确定MTHK通道的X射线结构。这与开放形式的MTHK的已知结构相结合，将为开放式和封闭构象中离子通道的详细原子分辨率图片提供第一个示例。第二个具体目的是在单克隆抗体的帮助下获得MTHK的高分辨率X射线结构。高分辨率结构将阐明配体结合位点特定相互作用的原子细节以及构象变化构成的蛋白质 - 蛋白质接触。第三个具体目的是研究Ca2+结合和通道门控之间耦合的功能力学。我们将使用基于结构的诱变与单个通道电生理记录相结合来分析配体门控的能量过程。